Project Summary/Abstract I have developed a career development strategy which balances course work in critical areas with intensive scientific development. My mentoring team is well-balanced by career stage and discipline. I will further develop my technical skills and scientific knowledge through their mentorship. Two of my mentors (Dr. Lee and Dr. Moses) engage in highly translational research. Additionally, enrollment in courses on grant writing and leadership training will enhance my ability to achieve and succeed in my career goal of becoming a highly successful independent researcher. If I am successful in my career transition to independence, I hope to use my training in glycoproteomics, small molecule quantitation, and quantitative proteomics to develop novel strategies to determine site-specific glycosylation pattern analysis. I hope to apply these strategies to the elucidation of the pathology and etiology of complex diseases involving glycosylation. The training I will receive through exposure to my mentors' scientific and translational expertise will be critical for my success as an independent researcher. The overall scientific goal of this proposal is to address critical challenges in the analysis of protein glycosylation, and determine protein glycosylation more thoroughly in the context of urinary tract infection, a condition known to be affected by glycosylation. Glycosylation is a critically important protein modification, and is understudied. This proposal investigates urinary protein glycosylation to new detail by performing a site- specific glycosylation profile (SSGP) analysis. Unfortunately, significant challenges remain preventing SSGP from becoming routine, and SSGP is currently low-throughput. We propose to improve SSGP analysis by developing a library of SSGP to make future SSGP analyses more accessible, faster and more sensitive. We will further use this library to establish the healthy baseline urinary SSGP among healthy individuals. In addition, glycosylation is known to affect urinary tract infections (UTI). Therefore a focused study will be performed to identify SSGP among individuals who have recurrent urinary tract infections, and to seek out new protective factors in urine. The aims of the study are: Aim 1: Determine urinary glycoprotein SSGP from patients with recurrent UTI and healthy controls; and Aim 2: Create a comprehensive library of SSGP in the urine of healthy individuals. I am extremely well supported in these scientific goals by an excellent research environment. Boston Children's Hospital (BCH) is consistently ranked among the finest children's hospitals in the US. There are 4 high-rise buildings housing researchers from BCH in the immediate vicinity of the hospital. There are dozens of internationally recognized scientists at BCH, including the primary mentor on this project, Dr. Marsha Moses. Two scientists have been recognized with the Nobel prize for contributions made at BCH. Importantly, there is an essential, well-categorized biobanking program within the department of Urology which will be essential to the success of this study.